Prophage WO genes recapitulate and enhance Wolbachia-induced cytoplasmic incompatibility

Abstract

The genus Wolbachia is an archetype of maternally inherited intracellular bacteria that infect the germline of numerous invertebrate species worldwide. They can selfishly alter arthropod sex ratios and reproductive strategies to increase the proportion of the infected matriline in the population. The most common reproductive manipulation is cytoplasmic incompatibility (CI), which results in embryonic lethality in crosses between infected males and uninfected females. Females infected with the same Wolbachia strain rescue this lethality. Despite more than 40 years of research1 and relevance to symbiont-induced speciation2,3, as well as control of arbovirus vectors4,5,6 and agricultural pests7, the bacterial genes underlying CI remain unknown. Here, we use comparative and transgenic approaches to demonstrate that two differentially transcribed, codiverging genes in the eukaryotic association module of prophage WO8 from Wolbachia strain wMel recapitulate and enhance CI. Dual expression in transgenic, uninfected males of Drosophila melanogaster crossed to uninfected females causes embryonic lethality. Each gene additively augments embryonic lethality in infected males crossed to uninfected females. Lethality associates with embryonic defects that parallel those of wild type CI and is notably rescued by wMel-infected embryos in all cases. The discovery of cytoplasmic incompatibility factor genes cifA and cifB pioneers genetic studies of prophage WO-induced reproductive manipulations and informs Wolbachia’s ongoing utility to control dengue and Zika transmission to humans.

Genome comparisons indicate recent transfer ofwRi‐likeWolbachiabetween sister speciesDrosophila suzukiiandD. subpulchrella

“…As noted by Beckmann and Fallon (2013), in w Ri, there is also a paralogous pair ( wRi_006720 and wRi_006710 ), termed “type II” by LePage et al. (2017), that exists within what they term a “WO‐like island.”…”

“…Sequences homologous to loci putatively involved in CI in other Wolbachia strains (Beckmann & Fallon, 2013; Beckmann et al., 2017; LePage et al., 2017) were extracted from w Ri (Klasson et al., 2009) and the draft assemblies for w Suz and w Spc. Differences among these three genomes at these loci were assessed by aligning the w Suz and w Spc reads to the w Ri reference and calculating the percentage of reads with the non‐ w Ri base.…”

“…We consider both a general pattern, namely the relative frequencies of nonsynonymous and synonymous substitutions, and sequence divergence for candidate loci associated with two Wolbachia ‐induced phenotypes, life shortening and CI. The “Octomom” duplication, which distinguishes w MelPop (Min & Benzer, 1997) from w Mel (Wu et al., 2004), contains the genes WD0507–WD0514 and is associated with extremely high Wolbachia titer and life shortening in D. melanogaster (Chrostek & Teixeira, 2015; but see Rohrscheib et al., 2016 for a critique and LePage et al., 2017 and Chrostek & Teixeira, 2017 for support of the hypothesis connecting these loci to life shortening or Wolbachia titer). Beckmann and Fallon (2013) used proteomics to identify the locus w Pip_0282 in w Pip, the Wolbachia found in Culex pipiens , as a candidate for producing CI.…”

“…Beckmann, Ronau, and Hochstrasser (2017) and LePage et al. (2017) provide experimental and bioinformatic evidence that WD0631 and WD0632 contribute to CI (but LePage et al. (2017) argue against the operon hypothesis).…”

“…(2017) provide experimental and bioinformatic evidence that WD0631 and WD0632 contribute to CI (but LePage et al. (2017) argue against the operon hypothesis). We examine differences in homologs and paralogs of these loci among w Suz, w Spc, and w Ri.…”